The present invention relates in general to scintillation spectral analysis of test samples containing n unknown isotopes (where n equals any whole integer) and, more particularly, to liquid scintillation spectral analysis techniques and equipment for analysis of samples containing n isotopes disposed in a liquid scintillator which are wholly automatic in operation and wherein the counting conditions are automatically optimized for each sample in response to determination of the effective quench level for each sample so that optimum counting conditions are attained irrespective of the actual quench level of each such sample. By the phrase "optimized counting conditions" it is meant that for multiple-labeled samples the ratio of the counting efficiency for each particular isotope to the counting efficiencies for all other isotopes in the sample is maximized in the counting window for that particular isotope while the counting efficiencies for the other isotopes are maintained constant in that particular window and, in the case of a single-labeled sample, the ratio E.sup.2 /B (where E is efficiency and B is background) is maximized, thus enhancing the statistical validity of a count in a given counting window for a given counting period. In its principal aspects, the invention is concerned with improved methods and apparatus for automatically adjusting, by predetermined preset amounts, the isotope counting window relative to the observed energy spectrum for any given isotope, such adjustment being made in response to detection of a measured parameter indicative of the effective quench level for each sample, whereby optimum counting conditions are created for each sample and for each isotope irrespective of the actual quench condition for each such sample.
Modern apparatus for detecting and measuring radioactivity has reached an unusually high state of development with systems currently available which offer unusual sensitivity to low energy radiation, as well as various options of full automation, semi-automation, or the more economical manual operating version. In a relatively few years, great strides have been made towards improving the preciseness and accuracy of counting efficiency in compliance with the very stringent requirements of users of this highly technical and sophisticated equipment. However, certain problems have continued to plague both the manufacturers and users of such equipment. A particularly prevalent and vexing problem has been the error introduced into computations of true sample activity levels because of a phenomenon commonly encountered with liquid scintillation samples known as "quenching". Stated very simply, this phenomenon results in attenuation of light scintillations within the samples, thus significantly affecting the statistical accuracy of the equipment which determined activity levels based upon the number and energy of such light scintillations, the latter being counted over known units of time and being proportional in energy to the energy of the disintegrations which produce them.
Many efforts have heretofore been made to minimize and, preferably, to eliminate, the errors which result from the quench phenomenon, some of which have completely failed and others of which have met with varying degrees of success and acceptance. One principal effort that has heretofore been made towards minimizing the quench problem has been that of development of various constituents which make up the sample and which are as free of quench characteristics as possible. Such constituents include, without limitation thereto, scintillation substances, solvents, and the material from which the light transmissive sample vial is made. However, perfect light transmitters completely devoid of quench characteristics are simply not available, and even if they were, the problem would remain since the test specimen itself may, and often will, contain quench materials such, for example, as blood or urine, which tend to attenuate the light because of their color. Moreover, unless the detection system is maintained in a completely enclosed atmosphere of an inert gas such as argon, quench can occur simply because of the presence of air.
Faced with the seeming impossibility of eliminating the quench phenomenon as a source of error, numerous efforts have been made to cope with the problem by providing methods and apparatus for compensating for such errors. Typical systems which are currently employed and which have found great acceptance today by people employing this sophisticated equipment include systems in which an external standard source which emits highly penetrating radiations is placed adjacent the sample in the detection chamber during a portion only of its overall counting cycle. Light scintillations occurring in the sample are then counted during at least two discrete intervals, during one of which the scintillations are created only by the isotope in the sample and during the other of which the scintillations are created by the composite effect of the isotope and the external standard. Suitable electronic equipment is provided for separating the pulses from the two sources on the basis of their different energy levels and, therefore, those which are counted primarily from the external standard provide a fairly accurate indication of the degree of quenching present in the sample since the counting efficiency for such external standard is known or can be readily ascertained by use of an unquenched standard sample. Typical systems of this type are described in detail in Lyle E. Packard U.S. Pat. No. 3,318,468, issued June 8, 1965, as well as in the aforesaid Cavanaugh application Ser. No. 541,721, filed Apr. 11, 1966, both of which are assigned to the assignee of the present invention.
Both of the aforementioned prior systems are of the type which are commonly referred to as "external standardization" systems and both represent basic and significant improvements over earlier known systems described therein, such as "internal standardization" and "channels ratio" systems. In effect, however, all of these prior systems have had certain aspects which are common to one another, a principal one of which is that the measured quench correlation parameter (e.g., "net or gross external standard count," "external standard ratio," "channels ratio," etc.) generally provides an indication of the degree of quench present in the sample, which indication must then be compared with a previously prepared quench correlation curve in order to determine the counting efficiency. Once knowing the counting efficiency, the counts per minute (cpm) measured for the isotope being analyzed can be divided by counting efficiency to determine activity level in disintegrations per minute (dpm). Unfortunately, however, the quench correlation curve itself differs widely from instrument to instrument, from isotope to isotope, from channel to channel, with sample volume, and with other variable conditions. Consequently, it has been necessary to prepare many of such curves, the preparation of each one of which has been time consuming, expensive, and subject to numerous human errors. Moreover, once the curves are prepared, it is necessary that the measured quench correlation data be compared with them in order to determine counting efficiency, thus introducing even further danger of human error.
Even more significant, however, has been the fact that while such a correlation curve can be prepared, it is only as accurate as the number of points which actually define the curve. It has been established that such points simply do not fall on a straight line, or even on a smoothly curved line -- quite to the contrary, the points will be non-uniformly distributed in an unpredictable random pattern which only generally defines the correlation curve. Consequently, even when the technician uses extreme care in his computations, he has been forced to extrapolate or interpolate between known points and, since the extrapolated or interpolated data can vary significantly from the actual data, the computed efficiency can still vary greatly from actual efficiency with maximum errors on the order of up to 10% and average errors on the order of up to 2% being common, dependent upon the number of differently quenched standard samples selected to prepare the quench correlation curve.
Errors of the foregoing magnitude were simply not acceptable to the highly trained technical personnel who use this general type of equipment. Indeed, such errors are highly objectionable, and the more so in view of the high state of sophistication that the overall art has reached. Indeed, these errors were tolerated only because the prior systems briefly described above, and described in considerably greater detail in the aforesaid Packard U.S. Pat. No. 3,188,468 and Cavanaugh application Ser. No. 541,721, represented the best available solutions to the problem at that time.
Continued efforts have, however, been made towards providing a more satisfactory solution to the problem. For example, it has been suggested that true activity level for a sample can be computed simply by dividing the measured variable quench correlation parameter (e.g., "external standard ratio," "net external standard count," "channels ratio," etc.) into the measured value in counts per minute (cpm) for the sample undergoing analysis. This suggestion, however, is not satisfactory for many reasons. For example, it assumes that the quench correlation curve is a straight line, which it is not. The fact that the quench correlation curve is not a straight line adds to the magnitude of such errors with the result that errors on the order of up to 25% have often been experienced and, indeed, on some occasions errors many times that magnitude have been encountered.
It has also been proposed that the problem can be resolved by servo-adjusting in any of various known manners, overall system gain and/or the high voltage supply so as to restore the measurable quench correlation parameter to a value indicative of an unquenched sample and, thereafter, analyzing the sample as if it were unquenched. Again, however, such a proposed "solution" is no solution at all since the gain correlation curves do not coincide with nor follow the quench correlation curves and, consequently, the magnitude of error can be and often will be, even greater than that experienced with the interpolation/extrapolation techniques referred to above.
Today, quite satisfactory liquid scintillation counting procedures and equipment have been devised which are capable of compensating for inaccuracies introduced by the quench phenomenon. Typical of these solutions are the procedures and equipment described in the aforesaid Packard application, Ser. No. 630,892, and Cavanaugh application, Ser. No. 630,891, both of which are assigned to the assignee of the present invention. Both of such prior applications describe, inter alia, procedures and equipment for imposing upon each sample a precisely controlled simulated quench condition which, when added to the actual quench condition for the sample, creates an accurately known effective quench level for which counting efficiencies for the isotope or isotopes being analyzed are known with a high degree of accuracy. However, effective as these solutions have been in enhancing the accuracy of the ultimate computation of sample activity level by permitting the technician to operate at a point or points where counting efficiencies are accurately known, nevertheless they still do not provide for optimum or near-optimum counting conditions and, therefore, optimized statistical validity.
Stated another way, in the case of multiple-labeled samples such as a dual-labeled sample containing tritium (.sup.3 H) and carbon-14 (.sup.14 C), assuming that the .sup.3 H and .sup.14 C counting windows are preset for optimum counting conditions for unquenched samples, it is known that as quenching progressively increases, the counting efficiency for tritium in both windows will be progressively degraded, while the counting efficiency for carbon in the .sup.3 H window will be progressively increased to a point and in the .sup.14 C window will be progressively degraded. Thus, isotope separation in the .sup.3 H window is progressively degraded, thereby degrading counting conditions and statistical counting validity, particularly in the case of samples having relatively low tritium activity levels. Similarly, in the case of a single-labeled sample, the counting efficiency will be progressively degraded with progressively increased quench, thus degrading the ratio E.sup.2 /B and, therefore, counting conditions. In either case, this results from the shift of the energy spectra for the isotopes being analyzed relative to their respective counting windows as defined by the pulse height analysis channels. While the systems described in the aforesaid Packard application Ser. No. 630,892 and Cavanaugh application Ser. No. 630,891 will permit a more accurate determination of counting efficiencies, the foregoing problem inherently causes the more accurately known efficiencies for single-labeled samples to be degraded and, for multiple-labeled samples, a similar degradation of isotope separation as samples are progressively more quenched. This result precludes counting at optimized counting conditions and, therefore, precludes attainment of optimum or near-optimum statistical validity. On the other hand, while the known systems for servo-adjusting gain or high voltage supply will, to a degree, improve the E.sup.2 /B ratio of single-labeled samples and isotope separation when dealing with multiple-labeled samples and thus improve statistical validity, they do so without any significant improvement in the accuracy of the computed sample activity level since they fail to compensate for the problem stemming from a shift of the measured parameter indicative of quench level from a point where counting efficiency is accurately known to a point where it is not.
It is a general aim of the present invention to provide an improved data analysis system which overcomes the foregoing disadvantages and which is characterized by its reliability, its rapidity of operation, its accuracy, and by its enhanced statistical validity characteristics. In this connection, it is an object of the invention to provide improved radioactivity spectrometry methods and apparatus which provide for automatic optimization of counting conditions for successive test samples irrespective of the degree of actual quench present in any given sample. While not so limited in its application, the invention will find especially advantageous use when the measured variable parameter indicative of quench conditions takes the form of net external standardization ratios, although it can also be used in connection with other measurable variable parameters which also provide an indication of the degree of quenching such, merely by way of example, as net external standard counts or net channels ratios.
Stated another way, it is a general aim of the present invention to provide improved radioactivity spectrometry methods and apparatus wherein the preset counting windows for the isotopes undergoing analysis are automatically readjusted relative to the observed energy spectra for successive samples by preset predetermined amounts selected in response to measurement of any suitable quench indicating parameter so that counting conditions are optimized automatically for each sample and for each isotope contained therein, irrespective of the actual quench level of each sample, all without requiring the attendance or attention of a technician during the analysis of successive samples having diverse quench levels on an automated basis.
As a consequence of attaining the foregoing general objectives of the invention, it has been found that when dealing with multiple-labeled samples considerably enhanced isotope separations are achieved and, when dealing with single-labeled samples, the E.sup.2 /B ratio is maximized, thereby insuring greater statistical validity.
In another of its important aspects, it is an object of the invention to provide improved methods and apparatus for optimizing counting conditions in radioactivity spectrometry applications characterized by their ability to permit highly accurate determination of sample activity levels under conditions of precisely known counting efficiencies irrespective of actual sample quench conditions, so that conditions of optimum accuracy and optimum statistical validity are achieved.
A more detailed object of the invention is the provision of improved methods and apparatus suitable for use in analysis of multiple-labeled samples wherein the ratio of the counting efficiency for each particular isotope to the counting efficiencies for all other isotopes in the sample is maximized in the counting window for that particular isotope while the counting efficiencies for the other isotopes are maintaned constant in that window, thus insuring attainment of optimized counting conditions. In this connection it is a related object of the invention to provide such improved methods and apparatus which will also find advantageous use in analysis of single-labeled samples by virtue of their ability to maximize the E.sup.2 /B ratio, whereby optimized counting conditions are attained.
A further and still more detailed objective of the invention is the provision of improved methods and apparatus of the character hereinabove set forth wherein provision is made for automatically readjusting by preset predetermined amounts selected window defining discriminator levels so that when analyzing multiple-labeled samples, the counting efficiencies for isotopes not of interest in any given counting window are maintained constant and, so that when analyzing single-labeled samples, the ratio E.sup.2 /B is maximized. In this connection it is a correlative object of the invention to provide improved methods and apparatus of the foregoing character wherein such results are achieved by automatically readjusting gain factors independently in the diverse analyzing channels by preset predetermined amounts so as to insure that the counting efficiencies for isotopes not of interest in any given window are maintained constant in the case of multiple-labeled samples and, in the case of single-labeled samples, the ratio E.sup.2 /B is maximized.
It is a more specific object of the invention to provide improved methods and apparatus for optimizing counting conditions automatically in response to measurement of a suitable quench indicating parameter, which measurement is also utilized to simulate a quench condition for any given sample so that such sample can be analyzed as if it were quenched to a level where counting efficiencies are precisely known, thus insuring not only optimum statistical validity, but also optimum computational accuracy .